The objective of this project is to uncover the molecular mechanisms of genetic rearrangements. The transposition reaction of bacteriophage Mu is studied as a model system. Critical steps in Mu transposition are a pair of DNA cleavages and strand transfers involving the ends of Mu DNA sequence and a target DNA; these reactions generate a branched DNA intermediate. These chemical reaction steps take place within a higher order protein-DNA complexe called transpososome, the core of which is composed of two Mu-end DNA segments synapsed by a stably bound tetramer of MuA transposase protein. Transpososome assembly normally is controlled by a number of cofactors: an enhancer- type DNA sequence element called IAS, the MuB protein, the E. coli- encoded HU and IHF proteins, ATP, and Mg++. The assembly process and the functional organization of the transpososome core have been studied by making use of a variety of mutant MuA proteins with missing functional domains. Structurally and functionally important protein-DNA interactions within the transpososome were analyzed by assembling it from short Mu end DNA fragments and MuA under permissive reaction conditions, bypassing the need for many of the cofactors normally required for the process. Both the Mu end DNA cleavage and the subsequent strand transfer at one Mu DNA end were shown to be catalyzed by the MuA monomers that were bound to the partner Mu DNA end within a transpososome; this explains why Mu DNA end synapsis is a prerequisite for the catalytic steps. The structure of the catalytic core domain of MuA has been determined by X-ray crystallography. The MuA catalytic subdomain shares remarkably similar structural arrangements with the core domain of HIV integrase which carries out similar biochemical reactions. The crystal structure of IHF in complex with its cognate DNA sequence has also been determined in collaboration with scientists in LMB/NIMH, demonstrating the DNA is bent into a U-shape by the bound protein. In collaboration with scientists in LCP/NIDDK, three independently folding DNA binding domains of MuA, one for the sequence at the IAS and two for the sites at the Mu ends, have been studied for their DNA binding characteristics, and their solution structures determined by NMR techniques.